Project Summary: Highly active antiretroviral therapy (HAART) has had a fundamental impact on the control of HIV-1 infection. HAART is remarkably effective at suppressing HIV-1 replication; however, it cannot completely eradicate the virus even during long-term therapy. A primary cause for the lack of complete viral clearance is a population of latently infected cells that appear to be a source for viral rebound upon withdrawal of antiretroviral therapy, thus perpetuating the infection and replenishing the latent reservoirs. Attempts to eliminate the latently infected cells by activating them with cytokines and lymphokines have not met with success probably because this treatment is unable to reach all of the sites that harbor latent virus and carries with it a high degree of toxicity. A promising alternative method using small molecules with pharmacological properties that would penetrate even the most inaccessible viral reservoirs and subsequently activate latent HIV-1 proviruses could very well improve the prospects for clearance of HIV-1. In this scenario, antiretroviral drugs, virus-induced cytotoxicity and immune-mediated elimination would control infection of freshly activated cells, while coincident administration of the anti- latency drug would not only force the emergence of latent virus but would also prevent the establishment of new latency. We developed a latency model that can be used for high throughput screening (HTS). With this system a very reliable signal was detected in 96- and 384-well formats as characterized by excellent Z' scores ranging between 0.55 and 0.80. Here we propose to utilize our model system for HTS to identify small molecule antagonists of latency. Once our library of low molecular weight molecules has been screened, the most active compounds will be further characterized both to probe the mechanism underlying the maintenance and reactivation of latent virus as well as providing molecules for further therapeutic development. We believe this system provides an efficient, rapid and safe method to discover novel agents ultimately aimed at elimination of HIV-1 infection. Relevance: When a person is infected with HIV, a small percentage of the cells that have been infected harbor viruses that can remain dormant for long periods of time, a phenomenon known as viral latency. Existing antiviral therapies are able to control actively replicating virus, but they cannot purge the hidden, latent virus, so patients remain perpetually infected because the latent virus may be activated at any time, leading to new rounds of viral production. Our goal is to identify new compounds that help to understand how to flush out the dormant virus and eliminate the infection in order to aid in reversing the course of the HIV pandemic. [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable]